Stable freeze-dried pharmaceutical formulation

ABSTRACT

The subject of the invention is a freeze-dried formulation consisting of an amorphous phase and a crystalline phase, which is pharmaceutically acceptable, comprising at least one nonprotein active ingredient, characterized in that it contains mannitol and alanine in a ratio R of between 0.1 and 1, R representing the mass of mannitol to the mass of alanine.

This application is a 371 of PCT/FR 96/01706 filed Oct. 30, 1996.

The present invention relates to a pharmaceutical formulation providedin the form of a freeze-dried product and containing at least one activeingredient of nonprotein nature. More particularly, the inventionrelates to such a formulation, stable at temperatures which may be ashigh as 25° C. to 40° C., which may be either reconstituted in liquidform by addition of a solvent for its administration via the parenteralor oral route, or directly administered via the oral route, to man or toanimals.

The active ingredient contained in the formulation according to theinvention may be alone or else combined with another active ingredientof protein or nonprotein nature.

It is known that freeze-drying may have a considerable effect on thedegradation of the pharmaceutical active ingredients in a formulation,as well as a strong impact on their stability in freeze-dried form. Thevarious variables which affect these parameters are mainly the pH, thequantity of salts present, the type and quantity of excipients in theformulation, the type of cryoprotection chosen, as well as thetemperatures, pressure and time chosen for the freezing, sublimation anddrying operations. These different variables influence the physicalstate of the freeze-dried product obtained, namely: vitreous amorphous,soft amorphous, crystalline or a combination of these states.

For the preservation of the freeze-dried products, amino acids,preferably glycine, and polyols, preferably mannitol, are often used;but the literature, which is highly abundant on the subject, gives noinformation on the solution to the general problem of obtaining a stablepharmaceutical formulation which takes into account the differentparameters involved in the operations for formulating and freeze-dryinga nonprotein active ingredient in combination with an amino acid and apolyol.

More particularly, the literature teaches that the presence of an aminoacid, of a polyol, for example mannitol, of a crystalline phase or of anamorphous phase may have, besides certain advantages, disadvantageswhich lead, in the case of freeze-dried products containing particularlysensitive active ingredients, to relatively short shelf lives and/orstorage temperatures for these freeze-dried products which are less than8° C. It would, however, be particularly advantageous, especially for anambulatory treatment, to be able to obtain a formulation which is stableat room temperature until it is reconstituted and to thereby avoid itsstorage in a refrigerator before or during treatment.

The role of the polyol and of amino acid has been studied separately inthe case of the human growth hormone (hGH), but their synergistic effectis still poorly elucidated (Pikal M. J., Dellermann K. M., Roy M. L.,Riggin M. N., The effects of formulation variables on the stability offreeze-dried Human Growth Hormone, Pharm. research., 1991, 8, No. 4,427-436).

The advantages and disadvantages linked to the presence of amino acids,of mannitol, of a crystalline phase or of an amorphous phase are listedbelow.

Advantages linked to the presence of amino acids.

It has been demonstrated that the presence of glycine in a freeze-driedproduct induced crystallization of the molecules present in solutionduring the freezing stage of the freeze-drying (Korey D. J., Schwartz J.B., Effects of excipients on the cristallization of pharmaceuticalcompounds during lyophylization, J. Parenteral Sci. Tech., 1989, 43, 2,80-83). This crystallization of the active ingredient makes it possibleto enhance its stability.

Alanine, in crystallized form, has the advantage of preventing thecollapse of the freeze-dried product during sublimation and drying andor allowing the production of a freeze-dried product with a greaterspecific surface area and therefore allows a more rapid drying (Pikal M.J., Freeze-drying of proteins, Biopharm., 26-30 October 1990).

Disadvantages linked to the presence of amino acids.

The addition of an amino acid to a sugar or to a polyol in a solution tobe freeze-dried generally has the effect of decreasing the glasstransition temperature of the sugar (te Booy M. P. W. M., de Ruiter R.A., de Meere A. L. J., Evaluation of the physical stability offreeze-dried sucrose containing formulations by differential scanningcalorimetry, Pharm. Research., 1992, 9, 109-114). Now, a decrease in theglass transition temperature is generally synonymous with a lowerstability of a freeze-dried product (Franks F., Freeze-drying; fromempiricism to predictability, Cryo-letters, 1990, 11, 93-110).

Advantages linked to the presence of mannitol.

The presence of mannitol in the composition of a freeze-dried product isgenerally justified as freeze-drying ballast, that is to say that itmakes it possible both to maintain the solid and rigid structure of thevolume of the freeze-dried product corresponding to the volume ofsolution to be freeze-dried, but its presence also makes it possible toadjust the isotonicity of the reconstituted solution to be injected.When mannitol is the predominant excipient in the composition of afreeze-dried product, it is most often in crystalline form (Lyophilizedformulations recombinant tumor necrosis factor, Hora M. S., Rana R. K.,Smith F. W., Pharm. Res., 1992, 9 (1), 33-36).

Disadvantages linked to the presence of mannitol.

It has been reported that the degree of hydrolysis of methylprednisolonesodium succinate, in freeze-dried form, was greater in the presence ofmannitol than in the presence of lactose, and that this level increasedwith the quantity of mannitol present in the freeze-dried product. Thishas been explained by the fact that the crystallization of mannitolduring freeze-drying changes the distribution of water in the matrix ofthe freeze-dried product. The increase in the quantity of water presentin the microenvironment of the active ingredient resulting therefromenhances the hydrolysis of the active ingredient and reduces itsstability (The effect of bulking agent on the solid state stability offreeze dried methylprednisolone sodium succinate, Herman B. D., SinclairB. D., Milton N., Nail S. L., Pharma. Res., 1994, 11 (10), 1467-1473).

Advantages linked to the presence of a crystalline phase.

The presence of a crystallized solute in a frozen solution is a means ofstabilizing the proteins during drying (Carpenter J. F. & Crowe J. H.,Modes of stabilization of a protein by organic solutes duringdessiccation, Cryobiology, 1988, 25, 459-470). Furthermore, thecrystallization, during freezing, of the predominant excipients presentin a solution to be freeze-dried makes the secondary sublimation anddrying operations more effective by increasing the specific surface areafor exchange between the atmosphere in the freeze-drying vessel and thesolid to be sublimed. This increase in the specific surface area of thecrystalline forms compared with the amorphous forms facilitates heatexchanges during freeze-drying. The consequence of this increasedefficiency in the freeze-drying is the production of freeze-dried formswhose residual water content is lower, which means an increasedstability of the freeze-dried product at higher temperatures (Korey D.J., Schwartz. J. B., Effects of excipients on the cristallization ofpharmaceutical compounds during lyophylization, J. Parenteral Sci.Tech., 1989, 43, 2, 80-83).

Disadvantages linked to the presence of a crystalline phase.

In general, the crystallized substances have less rapid dissolutionrates than the amorphous substances. Indeed, more energy is required todetach a molecule from an organized lattice of a crystalline arrangementthan to detach it from a disorganized assembly of an amorphous state.Sometimes, the dissolution rate becomes insufficient to allow asufficiently rapid absorption of these substances, which may lead to adecrease in their activity, especially in the case of molecules whichare not very stable in solution. Likewise, the perfect regularity ofcrystals being an ideal case, the heterogeneity of the crystalline phaseand the polymorphism which are obtained for the same substance andbetween associated substances induce different rates of dissolution forthe same substance and between each of the substances, which may resultin unreproducible therapeutic effects (Galénica 2, Biopharmacie 2ndedition, 1982, technique and documentation).

In addition, it has been demonstrated that the loss of activity of afreeze-dried protein was directly linked to the degree of crystallinityof the cryoprotective molecule (Izutsu K. L., Yoshioka S., Terao T.,Decreased protein-stabilizing effects of cryoprotectants due tocrystallization, Pharm. Research. 1993, 10, No. 8, 1232-1237; Izutsu K.I., Yoshioka S., Kojima S., Increased stabilizing effects of amphiphilicexcipients on freeze drying of lactate dehydrogenase (LDH) by dispersioninto sugar matrixes, Pharm. Res., 1995, 12 (6), 838-843). In theformulation of medicines containing proteins, the crystallization of theexcipients should be avoided according to: (Hermansky M., Pesak M.,Lyophilization of drugs, VI Amorphous and Cristalline forms Cesk. Farm.,1993, 42, (2), 95-98).

Advantages linked to the presence of an amorphous phase.

Based on the same line of thinking, the amorphous form dissolves morerapidly than the crystallized form and does not exhibit thedisadvantages linked to the heterogeneity and to the polymorphism of thecrystallized substances.

Moreover, the presence of additives in the amorphous state stabilizesthe activity of certain enzymes proportionally to the concentration ofthe additive according to Izutsu K. L., Yoshioka S., Terao T., Decreasedprotein-stabilizing effects of cryoprotectants due to crystallization,Pharm. Research., 1993, 10, No. 8, 1232-1237.

The cryoprotective effect of the excipients is attributed to theamorphous state of the glycine in the freeze-dried product obtained(Pikal M. J., Dellermann K. M., Roy M. L. Riggin M. N., The effects offormulation variables on the stability of freeze-dried Human GrowthHormone, Pharm. Research., 1991, 8, No. 4, 427-436).

Disadvantages linked to the presence of an amorphous phase.

In the presence of a solid amorphous phase alone, the freeze-driedproduct collapses at temperatures greater than the glass transitiontemperature during freezing. Within a soft amorphous phase, the chemicaldegradation reactions have much more rapid kinetics than within acrystalline phase (Solid state stability and preformulation study of anew parenteral cephalosporin antibiotics (E1040), Ashizawa K., UchikawaK., Hattori T., Ishibashi Y., Miyake Y., Sato T., Yakugaku Zasshi, 1990,110 (3), 191-201).

Furthermore, the higher rate of dissolution of the amorphous substancesis sometimes accompanied by a greater instability, the conversion of aform generally occurring from the amorphous state to the crystallizedstate (Galénica 2, Biopharmacie 2nd edition, 1982, technique etdocumentation).

In conclusion, the scientific literature on the subject of the effect ofexcipients on the stabilization of pharmaceutical active ingredientsgives contradictory information on their properties and furthermore doesnot make it possible to obtain some information on the subject of therelationships between the structure of a freeze-dried product and itsstability. Likewise, the role of the polyols and of the amino acids,alone or in combination, is not described according to a set ofgeneralizable properties, but has been observed with contradictoryresults according to the active principles studied and the quantities ofexcipients used.

It has now been found that a synergistic effect exists between mannitoland alanine on the stabilization of freeze-dried pharmaceutical activeingredients. It has in particular been demonstrated that thissynergistic effect exists only in a narrow range of relativeconcentrations of each of these two excipients.

The discovery of a surprising synergistic effect resulting from thecoexistence of an amorphous phase and a crystalline phase which has theconsequence of stabilizing the freeze-dried pharmaceutical activeingredient forms the basis of the present invention. The presentinvention therefore describes the production of this effect for specificmannitol/alanine ratios.

Thus, the present invention relates to a freeze-dried pharmaceuticalformulation consisting of an amorphous phase and a crystalline phase,comprising an effective quantity of at least one nonproteinpharmaceutical active ingredient, mannitol and alanine, the latter twoexcipients being in a mass ratio R of between 0.1 and 1, R being theratio of the mass of the mannitol to the mass of the alanine. The activeingredient included in the said formulation remains stable attemperatures which may range from 25° C. to 40° C. in freeze-dried form.Where appropriate, the dissolution of the freeze-dried product obtainedis rapid and complete. The freeze-dried product does not have acollapsed appearance and its water content is compatible withmaintaining the stability of the active ingredient.

It has been demonstrated that, for R of between 0.1 and 1:

the freeze-dried product consists of an amorphous phase and acrystalline phase,

the amorphous phase predominantly consists of mannitol and activeingredient,

the crystalline phase predominantly consists of alanine

Although the invention is not limited to a specific theory whichexplains the stabilization obtained by combining one or more nonproteinactive ingredients, mannitol and alanine in the indicated ratios, thefollowing hypothesis can be made:

the amorphous phase, demonstrated by differential scanning calorimetry,cryoprotects the pharmaceutical active ingredient during freezing, theactive ingredient itself being dispersed in this amorphous form, and thecrystalline phase, demonstrated by X-ray diffractometry, fixes thestructure of the freeze-dried product and avoids its collapse.

According to another of these features, the subject of the presentinvention is the production of stable freeze-dried products containing apharmaceutical active ingredient cryoprotected by an amorphous solidphase consisting completely or partially of mannitol, this amorphousphase coexisting within the freeze-dried product obtained aftersublimation and drying of the frozen solution, with a crystalline phaseconsisting essentially of alanine.

Thus, the subject of the present invention is also a process for thepreparation of freeze-dried pharmaceutical formulations comprising atleast one nonprotein active ingredient characterized in that a mixtureof the said active ingredient, mannitol and alanine in which themannitol and alanine are present in the ratio of between 0.1 and 1, Rbeing the ratio of the mannitol mass to the alanine mass,isfreeze-dried.

Other pharmaceutically acceptable excipients normally used infreeze-dried forms may be introduced into the formulation according tothe present invention, such as for example buffers or acid-bases whichmake it possible to adjust the pH, surfactants, salts, preservatives,especially antibacterial preservatives, antioxidants or chelatingagents, excluding excipients which, in the freeze-dried productcontaining the active ingredient, would prevent the coexistence of thecrystalline phase consisting predominantly of mannitol and of thecrystalline phase consisting predominantly of alanine, such as forexample certain protein derivatives of animal or plant origin such asgelatines, dextrins or proteins extracted from wheat grain or soya bean,gums such as agar or xanthan, polysaccharides, alginates,carboxymethylcelluloses, pectins, synthetic polymers such aspolyvinylpyrrolidone or complexes of a polysaccharide nature such asacacia gelatine. Among the buffers which may be introduced into theformulation according to the present invention, there may be mentionedin particular carbonate, borate, phosphate, citrate,tri(hydroxy-methyl)aminomethane, maleate and tartrate buffers, it beingunderstood that the acids and bases composing said buffers may also beintroduced alone. Among the surfactants which may be introduced into theformulation according to the present invention, there may be mentionedin particular polysorbates, poloxamers, tyloxapol, lecithins. Among thesalts which may be introduced into the formulation according to thepresent invention, there may be mentioned in particular the sodium saltssuch as ededate (tetrasodium EDTA), chloride, docusate (sodium1,4-bis(2-ethylhexyl)sulphosuccinate), bicarbonate, glutamate; potassiumacetate; dipotassium carbonate and magnesium stearate.

Among the preservatives which may be introduced into the formulationaccording to the present invention, there may be mentioned in particularmethyl and propyl para-hydroxybenzoate, benzethonium chloride, sodiummercurothiolate, phenylmercuric nitrate, benzyl alcohol, phenol andmetacresol.

The coexistence of the amorphous mannitol phase and the crystallinealanine phase is independent of the presence and of the concentration ofthe buffer used to adjust the pH of the solution, but it depends on theratio R defined above.

Examples of formulation of the solutions to be freeze-dried leading tothe formulations of the invention are the following:

One or a combination of pharmaceutical active ingredients, apharmaceutically acceptable buffer for adjusting the pH, mannitol andalanine with a mass ratio R=mass of mannitol/mass of alanine of between0.1 and 1, water for injectable preparations, as well as, if necessary,antibacterial preservatives and excipients which allow solubilization ofthe active ingredient or ingredients. According to a preferredembodiment of the invention, the alanine/mannitol mixture ispredominant.

The quantity of active ingredient present is limited by its solubilityin water. The formulations of the invention indeed result from thefreeze-drying of aqueous solutions in which the active ingredient isperfectly dissolved.

Likewise, any excipient is present in the formulation in a quantitybelow the quantity of the alanine/mannitol mixture.

The solutions to be freeze-dried are prepared in the following manner:

The desired quantities of buffer, alanine, mannitol, preservatives andactive ingredient are added, at the appropriate dissolution temperature,to the quantity of water for injectable preparations or of solubilizingagent necessary for their solubilization until complete dissolution isobtained. The solutions obtained are filtered in a sterile medium anddistributed into containers, preferably vials or capsules.

The freeze-drying of the solutions is carried out as follows:

The solution follows a cycle comprising freezing, then sublimation anddrying adapted to the volume to be freeze-dried and to the containercontaining the solution.

Preferably, a freezing rate close to −2° C./min is chosen in an Usifroid(France) freeze-drier type SMH15, SMJ100 or SMH2000.

The sublimation and drying times, temperatures and pressures areadjusted according to the volumes of solution to be freeze-dried and theresidual water content desired in the freeze-dried product.

A freeze-dried product is then obtained in which the alanine exists incrystallized form, and the mannitol in a completely or partiallyamorphous form. The freeze-dried product may be stored at 25° C. andeven up to 40° C. without adversely affecting the chemical andbiological stability of the active ingredient which it contains.

Full information on the techniques of preparation of injectableformulations by dissolution of the compositions of the invention isavailable to a person skilled in the art in Remington's PharmaceuticalSciences, 1985, 17th Edition or in William N. A. & Polli G. P., Thelyophilization of pharmaceuticals: a literature review, J. ParenteralSci. Tech., 1984, 38, (2), 48-59 or in Franks F., Freeze-drying: fromempiricism to predictability, Cryo-letters, 1990, 11, 93-110.

The active ingredient or the combined active ingredients, of thenonprotein type, formulated according to the present invention may beanalgesics, anti-inflammatory agents, antispasmodic agents, anticanceragents or active ingredients which can be used in cardiology, angiology,gastro-enterology, haematology and haemostasis, hepatology,infectiology, neurology-psychiatry, rhinology, rheumatology, toxicology,urology, or in the diagnostic field or as metabolism and nutritionregulators.

In the therapeutic families and the field of biological activity whichare mentioned above by way of example, any product can constitute theactive ingredient of the formulations of the present invention whichrepresent a considerable technical advance in the pharmaceuticaltechnique. Preferably, the active ingredients which are most adapted tothe formulations of the present invention are those whose stability inaqueous solution is problematical. It is however conceivable to applythe present invention to active ingredients which have no specificproblem of stability.

In the text which follows, the international nonproprietary names havebeen adopted to designate the active ingredients.

The active ingredient of the freeze-dried pharmaceutical formulations ofthe present invention may be chosen especially from the group consistingof:

phenylalkanoic acids, for example ketoprofen;

nonsteroid anti-inflammatory agents of the “oxicam” type, for examplepiroxicam, isoxicam, tenoxicam;

paracetamol;

lysine or arginine acetylsalicylate;

corticosteroids, for example methylprednisolone;

phloroglucinol;

bile acids, for example ursodeoxycholic acid or one of itspharmaceutically acceptable salts with inorganic or organic bases,preferably its sodium salt;

anthracyclines, for example doxorubicin, epirubicin, idarubicin,daunorubicin, pirarubicin;

platinum derivatives, for example cisplatin, oxaliplatin, carboplatin;

derivatives of alkaloids from Vinca minor, for example vinblastine,vincristine;

derivatives of alkaloids from rye ergot, for example dihydroergotamine,dihydroergotoxine, nicergoline;

derivatives of purine or pyrimidine bases, for example acyclovir,gancyclovir, cytarabine;

prostaglandins, for example sulprostone, alprostadil;

benzodiazepines, for example dipotassium clorazepate, devazepide;

beta-lactam antibiotics, for example piperacillin, tazobactam;

macrolide antibiotics, for example erythromycin or one of itsderivatives, in general a leukomycin;

antibiotics of the tetracycline family, for example minocycline;

antibiotics of the chloramphenicol type, for example thiamphenicol;

antibiotics of the spiramycin type;

nitrogenous mustards, for example chlorambucin and nitrosoureas, forexample carmustine and streptozocin. The nitrogenous mustards and thenitrosoureas are described in greater detail in Pharmacologie by M.Schorderet et al., 1992, 2nd edition, chapter 69, Ed., Frison, Roche,Paris;

H₂ antagonists, for example ranitidine, famotidine or one of theirpharmaceutically acceptable salts;

omeprazole and its analogues;

vitamins, for example thiamine, riboflavin, nicotinamide, pyridoxine,sodium panthotenate, biotin, ascorbic acid, folic acid, cyanocobalamin,retinol, cholecalciferol, alphatocopherol, cobalamide,hydroxycobalamide;

antitumour agents chosen from taxol, taxotere and their analogues,dacarbazine, methotrexate, plicamycin, thiotepa, streptozocin;

cardiovascular medicines chosen from molsidomine or one of itspharmaceutically acceptable salts, especially its hydrochloride,linsidomine, acetazolamide, meclofenoxate, diltiazem, sodiumnitroprussiate;

haematological medicines chosen from ticlopidine or one of itspharmaceutically acceptable salts, especially its hydrochloride,molgramostim, folinic acid;

anticoagulant and antithrombotic medicines chosen from heparin,low-molecular weight heparin in the form of nadroparin calcium,parnaparin sodium, dalteparin sodium, enoxaparin sodium, ardeparinsodium, certoparin sodium, reviparin sodium, minolteparin sodium,natural or synthetic antithrombotic pentasaccharides;

heparinoids, for example lomoparan;

diarginine oxoglutarate and the pharmaceutically acceptable salts ofoxoglutaric acid;

plant extracts, for example based on willow, harpagophytum, ginseng,fucus;

a gene, a DNA or RNA fragment intended for gene therapy, anoligonucleotide, an antisense oligonucleotide, nucleotides associatedwith protein compounds such as for example extracts of ribosomefractions, attenuated or inactivated live viruses;

valproic acid and its analogues;

metopimazine;

moxisylite;

pralidoxime;

deferoxamine;

phenobarbital or other barbiturates;

clomethiazole;

sodium pamidronate, sodium alandronate, sodium risendronate and otherdiphosphonates active as antiosteoporotic agent, especially{[(4-chlorophenyl)thio]methylene}bis(phosphonate) tiludronate ordisodium salt (SR 41319) in hemihydrate or monohydrate form;

5-HT₂ antagonists, especially ketanserine, ritanserine,(1Z,2E))-1-(2-fluorophenyl)-3-(4-hydroxyphenyl)-prop-2-en-1-one-O-(2-dimethylaminoethyl)oxime(SR 46349) or one of its pharmaceutically acceptable salts;

antagonists of angiotensin II, especially tasosartan, telmisartan,losartan potassium, losartan combined with hydrochlorothiazide (HCTZ),eprosartan, candesartan cilexetil, valsartan, irbesartan or2-n-butyl-3-{[2′-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl}-1,3-diazaspiro[4.4]non-1-en-4-one(SR 47436) and its pharmaceutically acceptable salts;

fantofarone or1-[(p-{3-[(3,4-dimethoxyphenethyl)methylamino]propoxy}phenyl)sulphonyl]-2-isopropylindolizine and its pharmaceutically acceptablesalts;

tirapazamine or 3-amino-1,2,4-benzotriazine-1,4-dioxide and itspharmaceutically acceptable salts;

(2S)-1-[(2R,3S)-5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzenesulphonyl)-3-hydroxy-2,3-dihydro-1H-indole-2-carbonyl]pyrrolidine-2-carboxamide(SR 49059) and its pharmaceutically acceptable salts;

N,N-dibutyl-3-{4-[(2-butyl-5-methylsulphonamido)-benzofuran-3-ylcarbonyl]phenoxyl}propylamineand its pharmaceutically acceptable salts, especially the hydrochloride(SR 33589);

6-(2-diethylamino-2-methyl)propylamino-3-phenyl-4-propylpyridazine (SR46559) and its pharmaceutically acceptable salts;

ethyl{(7S)-7-[(2R)-2-(3-chlorophenyl)-2-hydroxy-ethylamino]-5,6,7,8-tetrahydronaphthalen-2-yloxy}-acetateand its pharmaceutically acceptable salts, especially the hydrochloride(SR 58611A);

1-(2,4-dichlorophenyl)-3-(N-piperidin-1-yl-carboxamido)-4-methyl-5-(4-chlorophenyl)-1H-pyrazoleand its pharmaceutically acceptable salts, especially the hydrochloride(SR 141716A);

4-{[N-(3,4-dimethoxyphenethyl)]-N-methylamino-propoxyl}-2-benzenesulphonyl-3-isopropyl-1-methylindole(SR 33805) and its pharmaceutically acceptable salts;

2-{[1-(7-chloroquinolin-4-yl)-5-(2,6-dimethoxyphenyl)-1H-pyrazole-3-carbonyl]amino}adamantane-2-carboxylicacid (SR 48692) and its pharmaceutically acceptable salts;

N-cyclohexyl-N-ethyl-3-(3-chloro-4-cyclohexylphenyl)prop-2-enylamine (SR31747);

(−)-N-methyl-N-[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl)butyl]benzamide(SR 48968) and its pharmaceutically acceptable salts;

(S)-1-{2-[3-(3,4-dichlorophenyl)-1-(3-isopropoxyphenylacetyl)piperidin-3-yl]ethyl}-4-phenyl-1-azoniabicyclo[2.2.2]octanechloride (SR 140333A) and its pharmaceutically acceptable quaternarysalts, for example the benzenesulphonate;

4-amino-1-(6-chloropyrid-2-yl)piperidine and its pharmaceuticallyacceptable salts, especially the hydrochloride (SR 57227A);

(S)-N-(1-{3-[1-benzoyl-3-(3,4-dichlorophenyl)piperidin-3-yl]propyl}-4-phenylpiperidin-4-yl)N-methylacetamide(SR 142801) and its pharmaceutically acceptable salts;

2-{[4-(2-chlorophenyl) thiazol-2-yl]aminocarbonyl}-indole-1-acetic acid(SR 27897) and its pharmaceutically acceptable salts;

clopidogrel or methyl(+)-(S)-α-(2-chlorophenyl)-4,5,6,7-tetrahydrothieno-[3,2-c]pyridine-5(4H)-acetateand its pharmaceutically acceptable salts, especially its hydrogensulphate;

1-(2-naphthalen-2-ylethyl)-4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridine hydrochloride (SR 57746A) and its pharmaceuticallyacceptable salts, especially its hydrochloride;

N,N-dimethyl-N′-(pyridin-3-yl)methyl-N′-[4-(2,4,6-triisopropylphenyl)thiazol-2-yl]ethane-1,2-diamineand its pharmaceutically acceptable salts, especially the fumarate (SR27417);

2-[(5-(2,6-dimethoxyphenyl)-1-{4-[(3-dimethylaminopropyl)methylcarbamoyl]-2-isopropylphenyl}-1H-pyrazole-3-carbonyl)amino]adamantane-2-carboxylicacid and its pharmaceutically acceptable salts (SR 142948A);

3-(1-{2-[4-benzoyl-2-(3,4-difluorophenyl)morpholino-2-yl]ethyl}-4-phenylpiperidin-4-yl)-1,1-dimethylureaand its pharmaceutically acceptable salts (SR 144190A);

3-[N-{4-[4-(aminoiminomethyl)phenyl]-1,3-thiazol-2-yl}-N-(1-carboxymethylpiperidin-4-yl)amino]propionicacid trihydrochloride and its pharmaceutically acceptable salts (SR121566);

ethyl3-[N-{4-[4-(amino(N-ethoxycarbonylimino)methyl)phenyl]-1,3-thiazol-2-yl}-N-(1-ethoxy-carbonylmethyl)piperidin-4-yl)amino]propionate(SR 121787) and its pharmaceutically acceptable salts;

5-ethoxy-1-[4-(N-tert-butylcarbamoyl)-2-methoxybenzenesulphonyl]-3-spiro-[4-(2-morpholinoethyloxy)cyclohexane]indolin-2-one(SR 121463) and its pharmaceutically acceptable salts.

Most particularly preferred are the formulations of the invention inwhich the active ingredient is chosen from2-{[4-(2-enlorophenyl)thiazol-2-yl]aminocarbonyl}-indole-1-acetic acidor its potassium salt, irbesartan, clopidogrel, ursodeoxycholic acid andits sodium salt,1-(2-naphthalen-2-ylethyl)-4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridinehydrochloride,N,N-dimethyl-N′-(pyridin-3-yl)methyl-N′-[4-(2,4,6-triisopropylphenyl)thiazol-2-yl]ethane-1,2-diaminefumarate,2-[(5-(2,6-dimethoxyphenyl)-1-{4-[(3-dimethylaminopropyl)methylcarbamoyl]-2-isopropylphenyl}-1H-pyrazole-3-carbonyl)amino]adamantane-2-carboxylicacid,3-(1-(2-[4-benzoyl-2-(3,4-difluorophenyl)morpholino-2-yl]ethyl}-4-phenylpiperidin-4-yl)-1,1-dimethylurea,3-(N-{4-[4-(aminoiminomethyl)phenyl]-1,3-thiazol-2-yl}-N-(1-carboxymethylpiperidin-4-yl)amino]propionicacid trihydrochloride, ethyl3-[N-{4-[4-(amino(N-ethoxycarbonylimino)methyl)phenyl]-1,3-thiazol-2-yl}-N-(1-(ethoxycarbonylmethyl)piperidin-4-yl)amino]-propionate,5-ethoxy-1-[4-(N-tert-butylcarbamoyl)-2-methoxybenzenesulphonyl]-3-spiro-[4-(2-morpholinoethyloxy)cyclohexane]indolin-2-oneand their pharmaceutically acceptable salts.

The following formulations are particularly advantageous:

any formulation obtained by freeze-drying a solution in which themannitol is at a concentration of 9 mg per ml, the alanine is at aconcentration of 18 mg per ml and the active ingredient is2-{[4-(2-chlorophenyl)thiazol-2-yl]aminocarbonyl}indole-1-acetic acid ata concentration of 1.18 mg per ml or one of its pharmaceuticallyacceptable salts at an equivalent concentration;

any formulation obtained by freeze-drying a solution in which themannitol is at a concentration of 10 mg per ml, the alanine is at aconcentration of 23 mg per ml and the active ingredient is irbesartan ata concentration of 1 mg per ml or one of its pharmaceutically acceptablesalts at an equivalent concentration; and

any formulation obtained by freeze-drying a solution in which themannitol is at a concentration of 9 mg per ml, the alanine is at aconcentration of 18 mg per ml and the active ingredient is1-(2-naphthalen-2-ylethyl)-4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridinehydrochloride at a concentration of between 0.01 mg and 0.2 mg per ml orone of its pharmaceutically acceptable salts at an equivalentconcentration.

A pharmaceutically acceptable salt of any one of the salifiable activeingredients listed above may also be selected as active ingredient.

The pharmaceutical active ingredient is preferably chosen from the groupconsisting of the potassium salt of the acid SR 27897 designated belowas SR 27897B, irbesartan or SR 47436, clopidogrel, ursodeoxycholic acidor its sodium salt, SR 57746A and SR 27417A.

To illustrate the present invention, without however limiting it,evaluations were carried out by choosing, as an example ofpharmaceutical active ingredient, SR 27897B, SR 47436 (irbesartan) andSR 57746A. Thus, several solutions containing SR 27897B at 1 mg/ml, aphosphate buffer (Na₂HPO₄/NaH₂PO₄) at various molarities and at pHvalues of between 7.5 and 8.25; mannitol and alanine in a ratio R=massof mannitol/mass of alanine of between 0.1 and 1 were prepared,freeze-dried and analyzed.

Likewise, several solutions containing SR 47436 at 1 mg/ml, potassiumhydroxide in a molar ratio [KOH]/[SR 47436] greater than or equal to 1,alanine alone or a mannitol-alanine mixture in a ratio R=mass ofmannitol/mass of alanine of between 0.1 and 1, and ethanol, wereprepared, freeze-dried and analyzed.

Finally, a solution containing SR 57746A in hydrochloride form at 0.11mg/ml, anhydrous citric acid and a mannitol-alanine mixture in a ratioR=mass of mannitol/mass of alanine equal to 0.5 was prepared,freeze-dried and analyzed.

TABLE 1 below indicates the compositions of the solutions studiedcontaining SR 27897B. For each of these formulations, R=0.5 with, asmannitol, alanine and SR 27897B concentration, 9 mg/ml, 18 mg/ml and 1mg/ml respectively.

TABLE 1 SODIUM PHOSPHATE pH OF THE SODIUM BATCH NO. BUFFER: mM PHOSPHATEBUFFER  1  5 7.5  2  5 8  3 10 8  4 25 8  5 25 8.5  6 15 8  7 25 7.75  825 8  9 25 8.25 10 35 8.25 11 25 8

TABLE 2 below indicates the composition of the freeze-dried solutionsstudied containing SR 47436.

TABLE 2 BATCH mg/ml mg/ml mg/ml mg/ml NO. MANNITOL ALANINE R KOH SR47436 12 10 18 0.55 0.137 1 13 10 23 0.43 0.137 1

TABLE 3 below indicates the composition of the freeze-dried solutionsstudied containing SR 57746A in hydrochloride form.

TABLE 3 mg/ml BATCH mg/ml mg/ml ANHYDROUS mg/ml mg/m/ NO. MANNITOLALANINE R CITRIC ACID SR 57746A POLYSORBATE 80 14 9 18 0.5 7.7 0.11 0 159 18 0.5 7.7 0.11 1

The turbidity of the freeze-dried products taken up in solution will bedetermined with the aid of a Ratio Hach 18900-00 turbidimeter. Theresults will be expressed in nephelometric turbidity units (NTU) definedby Standard methods for the examination of water and waste-water of theAmerican Public Health Association.

The organoleptic criteria for the freeze-dried products will be examinedvisually and will take into account the colour of the freeze-driedproduct, its structure (collapsed or otherwise), as well as theobservation of a possible phase shift between the crust and the crumb ofthe freeze-dried product.

The water content of the freeze-dried products will be determined bycoulometry according to the method described in Ph. Fr. Xth Ed. V.3.5.6. A., by injecting 2 ml of methanol into the vial of freeze-driedproduct with the aid of a syringe. The water content will be expressedin percentage by weight of the freeze-dried product.

X-ray diffractometric analysis on the freeze-dried products will becarried out on a SIEMENS D500 TT diffractometer; source: CuKal;generator: 40 KV, 25 mA; back monochromator; slits: 1/1/1/0.16/0.6;sampling on a pyrex support; scanning range: 40 to 400 per minute forBragg 2 theta.

Differential scanning calorimetry (DSC) will be carried out using thePerkin Elmer DSC 7 apparatus with the following characteristics:calibration with indium and lead, sampling between 5 and 10 mg in a 50μl capsule, initial temperature of 10° C., heating rate of 10°C./minute, final temperature 300° C.

The SR 27897B assay will be carried out by liquid chromatography (Ph.Eur. 2 (1) V. 6.20.4. ) at 254 nm using a C18 graft column 25 cm long,4.6 mm internal diameter and particle size 10 μm (Bischoff reference25461840). The mobile phase will consist of a volume for volume mixtureof acetate buffer pH 4.0 (glacial acetic acid and concentrated ammoniumhydroxide Merck) and of acetonitrile for chromatography (Sharlaureference Ac33). The control solution will consist of a solution of SR27897B (provided by Sanofi Recherche) at 50 μg per ml of methanol (Merckreference 6009). The solution to be analyzed will be obtained bydissolving the freeze-dried product in 100 ml of ultrapurified water(Millipore, “Milli-Q” water). The flow rate will be 2 ml/min. Thespecific peak surface area obtained after injection of 20 μl of controlsolution and then of solution to be analyzed will be calculated for eachof the chromatograms. The SR 27897B content of the freeze-dried product,expressed in mg/vial, will be determined from the calculation of thesetwo surface areas.

The assay of the related substances (impurities) of SR 27897B in thefreeze-dried product during preservation, a parameter indicative of thestability of the product, will also be carried out by liquidchromatography on a C18 graft column (Bischoff reference 25461840). Themobile phase will consist of a gradient of acetonitrile and acetatebuffer pH 4.0 whose composition is indicated in TABLE A:

TABLE A ACETONITRILE ACETATE BUFFER pH 4.0 TIME (minute) (volume)(volume)  0 20 80  5 30 70 15 60 40 25 70 30 28 20 80 40 20 80

The control solution will consist of a solution of SR 27897B (SanofiRecherche) at 10 μg per ml of methanol. The solution to be analyzed willbe obtained by dissolving the contents of a freeze-dried vial in 5 ml ofmethanol. The flow rate will be 2 ml/min. The specific peak surface areaof the unknown impurities obtained on the chromatograms after injectionof 20 μl of the solution to be analyzed, expressed in terms of thespecific peak surface area of SR 27897B obtained after injection of 20μl of the control solution, will be calculated in the same manner. Thecontent of each of the unknown impurities and the overall content ofimpurities of the freeze-dried SR 27897B, expressed in percentage byweight of the product, can be determined from these calculations.

The SR 47436 assay will be carried out by HPLC liquid chromatography(Ph. Eur. 2 (1) V 6.20.4.) at 220 nm using a C18 graft silica columnmade of stainless steel, 25 cm long, 8 mm external diameter and 4 mminternal diameter, spherical silica of diameter 7 μm and of 120 Å porediameter having undergone an “end capping” treatment (reference column720042 supplied by Chromoptic). The mobile phase will consist of amixture of 60 volumes of phosphate buffer solution pH 3.0 (phosphoricacid Prolabo reference 20624295, triethylamine Fluka reference 90340)and of 40 volumes of acetonitrile for chromatography (Merck reference14291) with a flow rate of 1 ml/min.

The first control solution will consist of a solution of SR 47436(Sanofi Recherche) at 0.5 mg per ml of mobile phase. The second controlsolution will consist of a solution containing 0.5 mg of SR 47436 and0.5 mg of impurity corresponding to the opening product (SanofiRecherche) per ml of mobile phase. The solution to be analyzed will beobtained by dissolving the freeze-dried product in 10 ml of mobilephase. It will be checked by successive injection of the first andsecond control solutions that the operating conditions are satisfactory(resolution factor greater than 2 between the two peaks for an injectionof 10 μl of the second control solution, coefficient of variation of thesurface area of the peak less than or equal to 1% for a series of 5injections of 10 μl of the first control solution). After injection of10 μl of each control solution and of 20 μl of each solution to beanalyzed, the SR 47436 content in mg per freeze-dried product will bedetermined by calculating the specific peak surface areas obtained onthe chromatograms.

The assay of the related substances (impurities) of SR 47436 will becarried out by HPLC liquid chromatography (Ph. Eur. 2 (1) V 6.20.4.) at220 nm using a C18 graft silica column (cf. assay of SR 47436). Themobile phase will consist of a mixture of 60 volumes of phosphate bufferpH 3.1 and 40 volumes of acetonitrile for chromatography with a flowrate of 1 ml/min. The two control solutions will consist, for the first,of a solution of SR 47436 (Sanofi Recherche) at 0.5 mg per ml ofmethanol (supplied by SDS under the reference 0930221) and for thesecond, of a solution of SR 47436 at 0.5 μg per ml of methanol. Thesolution to be analyzed will be obtained by dissolving the freeze-driedproduct in 10 ml of water for injectable preparations (IP). The analysisshould be carried out within half an hour at the latest after thereconstitution. The existence of satisfactory operating conditions willbe checked by successive injections of 10 μl of water for injectablepreparations and 10 μl of the two control solutions (retention time ofthe principle peak similar for the two controls, signal to noise ratiogreater than or equal to 10 for the first control). After injecting 10μl of the solution to be analyzed, the content per related substance andthe overall content of related substances (impurities) expressed inpercentage of product weight will be determined by calculating thespecific peak surface areas obtained on the chromatograms.

The SR 57746A assay (Sanofi Recherche) will be carried out by liquidchromatography at 224 nm using a C18 graft silica column 25 cm long, 4mm internal diameter and particle size 7 μm (Macherey Nagel, reference720042). The mobile phase will consist of a mixture of 45 volumes ofacetonitrile for chromatography (Rathburn reference RH 1016) and 55volumes of buffer solution pH 3.0 (prepared by diluting 5.5 ml ofphosphoric acid in 950 ml of filtered demineralized water (MilliporeAlpha-Q), then by adjusting the pH to 3.0 with a triethylamine solution(Fluka, reference 90340) then by adding 10 ml of acetonitrile and byfilling to 1000 ml with filtered demineralized water). The controlsolution will consist of an SR 57746A solution at 15.0 mg per 100 ml ofmethanol (Carlo Erba, reference 414814). The solution to be analyzedwill be obtained by dissolving the freeze-dried product in 3.0 ml of amixture consisting of 25 volumes of methanol and 75 volumes of filtereddemineralized water. The flow rate will be 1 ml per minute. The specificpeak surface area obtained after injection of 10 μl of control solutionand then of solution to be analyzed will be measured for each of thechromatograms. The SR 57746A content of the freeze-dried product,expressed in mg/vial, can be determined from the measurement of the twosurface areas.

The assay of the related substances (impurities) of SR 57746A in thefreeze-dried product during preservation will be also carried out byliquid column chromatography with the chromatographic conditionsdescribed in “Assays” in (Ph. Eur. 2 (1) V.6.20.4. ). The controlsolution will consist of a solution of SR 57746A at 0.15 μg per ml ofmethanol. The solution to be analyzed will be obtained by dissolving thecontents of the freeze-dried product in 3 ml of a mixture of 25 volumesof methanol and of 75 volumes of filtered demineralized water. The flowrate will be 1 ml per minute. The specific peak surface area of theunknown impurities obtained on the chromatograms after injection of 10μl of the solution to be analyzed, expressed in terms of the specificpeak surface area of SR 57746A obtained after injection of 10 μl ofcontrol solution will be measured in the same way. The content of eachof the unknown impurities and the overall content of impurities of thefreeze-dried SR 57746A, expressed as percentage in terms of surfacearea, can be determined from these measurements.

The analytical results obtained using these various methods aredescribed below.

TABLE 4 below represents the results of the initial controls carried outon the freeze-dried products of SR 27897B for the water content (in % byweight of the freeze-dried product), the glass transition temperature Tg(in ° C. ) determined by DSC, and on the freeze-dried products taken upin water for IP for the turbidity (in NTU) and the pH.

TABLE 4 Batch No. % Water Tg ° C. Turbidity NTU pH  1 0.39 27 58 7.25  20.47 25 23 7.2  3 0.6 27 11 7.4  4 0.91 35 2.2 7.6  5 0.89 40 29 7.5  60.61 36.7 17 7.6  7 0.93 45.5 33 7.6  8 1.07 46.1 1.4 7.8  9 1.04 45.70.5 7.7 10 1.94 45.8 0.8 7.8 11 4.5 7.6

By way of an additional example, several batches of freeze-dried productof SR 27897B were monitored for stability at 50° C., 25° C., 40° C. and50° C. for 1 month, 3 months and 6 months.

TABLE 5 which represents the total content of related substances(impurities), expressed in % by weight of initial SR 27897B, found inthe freeze-dried products of SR 27897B after 1 month of storage showsthat the stability is excellent after this period of storage.

TABLE 5 Batch % Impurities % Impurities % Impurities % Impurities No. at5° C. at 25° C. at 40° C. at 50° C.  1 <0.27 <0.26 <0.25  2 <0.23 <0.26<0.24  3 <0.23 <0.26 <0.25  4 <0.26 <0.24 <0.24  5 <0.23 <0.25 <0.26  6<0.1  7 <0.1  8 <0.1  9 <0.1 10 <0.1 11 <0.1 <0.1

TABLE 6 which represents the total content of related substances(impurities) found in the freeze-dried products of SR 27897B after 3months of storage at 50° C., shows that the stability is excellent afterthis period of storage.

TABLE 6 % Impurities Batch No. at 50° C. 1 <0.1 2 <0.1 3 <0.1 4 <0.1 5<0.1

Finally, TABLE 7 represents the total content of related substances(impurities) found in the freeze-dried products of SR 27897B after 6months of storage at 5° C. and 40° C., shows that the stability is alsoexcellent after this period of storage.

TABLE 7 % Impurities % Impurities Batch No. at 5° C. at 40° C.  7 <0.10.13  8 <0.1 0.1  9 <0.1 0.1 11 <0.1

X-ray diffraction.

The result of the X-ray diffraction analysis on the powder of twofreeze-dried products containing a mannitol/alanine mixture in a ratioR=mass of mannitol/mass of alanine=0.5 is indicated in FIG. 1,diffractograms 1 and 2. The diffractograms 3 and 4 of FIG. 1 representthe controls alanine and mannitol. As can be observed in this figure,the line situated between 10° and 11°, characteristic of crystallizedmannitol, is not obtained for the two freeze-dried products of SR27897B. Thus, for R=0.5, the alanine alone is in crystallized form, themannitol being, for its part, in amorphous form.

Differential scanning calorimetry.

FIG. 2 represents the influence of the alanine to mannitol mass ratio onthe glass transition temperature of the freeze-dried products. Thisfigure shows us that the maximum glass transition temperature isobtained for (1/R)>1, that is to say for R of between 0 and 1. Ingeneral, the glass transition temperature is representative of themaximum temperature for stability of the freeze-dried product. Thus, themaximum temperature for stability of the freeze-dried product is reachedfor R of between 0 and 1.

TABLE 8 below represents the results of the initial controls carried outon the freeze-dried products of SR 47436 for the water content, thetotal content of related substances (impurities) and on the freeze-driedproducts taken up in water for IP, for the pH.

TABLE 8 Batch No. % Water % Impurities pH 12 0.5% 0.24 6.6 13 0.2 0.16.7

TABLE 9 below represents the total content of related substancesexpressed as percentage purity in terms of SR 47436 of the freeze-driedproducts of SR 47436 batch 12 after 1 week and 2 weeks of storage at 5°C., 25° C., 35° C. and 50° C.

TABLE 9 Batch No. 12 5° C. 25° C. 35° C. 50° C. 1 week  99.89 99.8999.71 98.47 2 weeks 99.90 99.82 99.47 97.05

TABLE 10 below represents the total contents of related substancesexpressed as percentage of impurities, of the freeze-dried products ofSR 47436 batch 13 after 3 months, 6 months and 9 months of storage at 5°C., 25° C. and 35° C.

TABLE 10 Batch No. 13 5° C. 25° C. 35° C. 3 months 0.1% 0.2% 0.3% 6months 0.2% 0.3% 0.6% 9 months 0.3% 0.3% —

TABLE 11 below represents the total contents of related substancesexpressed as percentage of impurities, of the freeze-dried products ofSR 57746A after 1 month and 3 months of storage at 5° C., 25° C. and 40°C. and of a freeze-dried product of SR 57746A reconstituted immediately(reference).

TABLE 11 Batches N^(o) 14 and 15 5° C. 25° C. 40° C. Reference <0.1% 1month <0.1% <0.1% <0.1% 3 months <0.1% <0.1% <0.1%

EXAMPLE 1

Composition of a freeze-dried product of SR 27897 (base) to be taken upin 1 ml of water for IP.

CONSTITUENTS Unit formula in (mg) SR 27897B* 1.18 mg Apyrogenic alanine18.0 mg Mannitol 9.0 mg Apyrogenic monosodium phosphate 0.3 mg dihydrateApyrogenic disodium phosphate 8.5 mg dodecahydrate White glass vial type1 of 3 ml 1 Grey chlorobutyl pillar stopper 1 Blue flip-off aluminiumcapsule 1 diameter 13 mm *corresponding to 1 mg of SR 27897 acid

EXAMPLE 2

Composition of a freeze-dried product of SR 27897 (base) to be taken upin 5 ml of water for IP.

CONSTITUENTS Unit formula in (mg) SR 27897B* 5.9 mg Apyrogenic alanine90.0 mg Apyrogenic mannitol 45.0 mg Apyrogenic monosodium phosphate 1.5mg dihydrate Apyrogenic disodium phosphate 42.5 mg dodecahydrate Whiteglass vial type 1 of 1 20 ml Grey chlorobutyl pillar stopper 1 diameter20 mm Blue flip-off aluminium capuule 1 diameter 20 mm *corresponding to5 mg of SR 27897 acid

EXAMPLE 3

Composition of a freeze-dried product of SR 47436 at 5 mg to be taken upin 5 ml of water for IP.

CONSTITUENTS Unit formula in (mg) SR 47436 5.0 mg Apyrogenic alanine115.0 mg Apyrogenic mannitol 50.0 mg Potassium hydroxide 0.687 mg Whiteglass vial type 1 of 1 20 ml Grey chlorobutyl pillar stopper 1 diameter20 mm Aluminium capsule with 1 operculum, diameter 20 mm

EXAMPLE 4

Composition of a freeze-dried product of SR 57746A (hydrochloride) to betaken up in 4 ml of water for IP.

CONSTITUENTS Unit formula in (mg) SR 57746A 0.44 mg Apyrogenic alanine72.0 mg Apyrogenic mannitol 36.0 mg Apyrogenic anhydrous citric 30.8 mgacid White glass vial type 1 of 1 20 ml Grey chlorobutyl pillar stopper1 diameter 20 mm Blue flip-off aluminium capsule 1 diameter 20 mm

EXAMPLE 5

Composition of a solution of SR 57746A (hydrochloride) to befreeze-dried, expressed in concentration for final volumes of solutionwhich may be as high as 100 ml by addition of a sufficient quantity ofwater for IP.

Unit formula CONSTITUENTS expressed in mg/ml SR 57746A 0.11 mg/mlApyrogenic alanine 18.0 mg/ml Apyrogenic mannitol 9.0 mg/ml apyrogenicanhydrous citric acid 7.7 mg/ml Water for injectable preparations qs 1ml White glass vial type 1 1 Grey chlorobutyl pillar stopper 1 Blueflip-off aluminium capsule 1

EXAMPLE 6

Composition of a freeze-dried product of SR 57746A (hydrochloride)containing from 0.01 mg to 0.2 mg of SR 57746A (hydrochloride) to betaken up in 1 ml of water for IP.

CONSTITUENTS Unit formula in (mg) Apyrogenic alanine 18.0 mg Apyrogenicmannitol 9.0 mg Apyrogenic anhydrous citric acid 7.7 mg White glass vialtype 1 of 3 ml 1 Grey chlorobutyl pillar stopper 1 Blue flip-offaluminium capsule 1 diameter 13 mm

EXAMPLE 7

Composition of a freeze-dried product of SR 57746A (hydrochloride) to betaken up in 4 ml of water for IP.

CONSTITUENTS Unit formula in (mg) SR 57746A 0.44 mg Apyrogenic alanine72.0 mg Apyrogenic mannitol 36.0 mg Apyrogenic anhydrous citric acid30.8 mg Polysorbate 80 4.0 mg White glass vial type 1 of 20 ml 1 Greychlorobutyl pillar stopper 1 diameter 20 mm Blue flip-off aluminiumcapsule 1 diameter 20 mm

EXAMPLE 8

Composition of a solution of SR 57746A (hydrochloride) to befreeze-dried, expressed in concentration for final volumes of solutionwhich may be as high as 100 ml by addition of a sufficient quantity ofwater for IP.

Unit formula CONSTITUENTS expressed in mg/ml SR 57746A 0.11 mg/mlApyrogenic alanine 18.0 mg/ml Apyrogenic mannitol 9.0 mg/ml apyrogenicanhydrous citric acid 7.7 mg/ml Polysorbate 80 1.0 mg/ml Water forinjectable preparations qs 1 ml White glass vial type 1 1 Greychlorobutyl pillar stopper 1 Blue flip-off aluminium capsule 1

EXAMPLE 9

Composition of a freeze-dried product of SR 57746A (hydrochloride)containing from 0.01 mg to 0.2 mg of SR 57746A (hydrochloride) to betaken up in 1 ml of water for IP.

CONSTITUENTS Unit formula in (mg) Apyrogenic alanine 18.0 mg Apyrogenicmannitol 9.0 mg Apyrogenic anhydrous citric acid 7.7 mg Polysorbate 801.0 mg White glass vial type 1 of 3 ml 1 Grey chlorobutyl pillar stopper1 Blue flip-off aluminium capsule 1 diameter 13 mm

What is claimed is:
 1. Freeze-dried formulation consisting of anamorphous phase and a crystalline phase, which is pharmaceuticllyacceptable, comprising at least one nonprotein active ingredient,characterized in that it contains mannitol and alanine in a ratio R ofbetween 0.1 and 1, R representing the mass of mannitol to the mass ofalanine, it being understood that formulations further comprising one ormore matrix forming agent selected from the group consisting of pectins,gelatins, soy fiber proteins and mixtures thereof are excluded. 2.Formulation according to claim 1, in which the active ingredient iscombined with another active ingredient of a protein nature. 3.Formulation according to claim 1 or 2, comprising, in addition, at leastone additional compound selected from the group consisting of a buffer,a surfactant, a preservative, a salt, an antioxidant and a chelatingagent.
 4. Formulation according to claim 1 or 2, for the reconstitutionof a solution for its administration via the parenteral route. 5.Formulation according to claim 1 or 2, for the reconstitution of asolution for its administration via the oral route.
 6. Formulationaccording to claim 4, for the reconstitution of an injectable solution.7. Formulation according to claim 1, which is directly administrable viathe oral route.
 8. Formulation according to claim 1, in which the activeingredient is chosen from2-{[4-(2-chlorophenyl)thiazol-2-yl]aminocarbonyl}indole-1-acetic acid orits potassium salt, irbesartan, clopidogrel, ursodeoxycholic acid andits sodium salt,1-(2-naphthalen-2-ylethyl)-4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridinehydrochloride,N,N-dimethyl-N′-(pyridin-3-yl)methyl-N′-[4-(2,4,6-triisopropylphenyl)thiazol-2-yl]ethane-1,2-diaminefumarate,2-[(5-(2,6-dimethoxyphenyl)-1-{4-[(3-dimethylaminopropyl)methylcarbamoyl]-2-isopropylphenyl}-1H-pyrazole-3-carbonyl)amino]adamantane-2-carboxylicacid,3-(1-{2-[4-benzoyl-2-(3,4-difluorophenyl)morpholino-2-yl]ethyl}-4-phenylpiperidin-4-yl)-1,1-dimethylurea,3-[N-{4-[4-(aminoiminomethyl)phenyl]-1,3-thiazol-2-yl}-N-(1-carboxymethylpiperidin-4-yl)-amino]propionicacid trihydrochloride, ethyl 3-[N-{4-[4-(amino (N-ethoxycarbonylimino)methyl)phenyl]-1,3-thiazol-2-yl}-N-(1-(ethoxycarbonylmethyl)piperidin-4-yl)amino]propionate,5-ethoxy-1-[4-(N-tert-butyl-carbamoyl)-2-methoxybenzenesulphonyl]-3-spiro-[4-(2-morpholinoethyloxy)cyclohexane]indolin-2-oneand their pharmaceutically acceptable salts.
 9. Formulation according toclaim 1, obtained after freeze-drying a solution in which the mannitolis at a concentration of 9 mg per ml, the alanine is at a concentrationof 18 mg per ml and the active ingredient is2-{[4-(2-chlorophenyl)thiazol-2-yl]aminocarbonyl}-indole-1-acetic acidat a concentration of 1.18 mg per ml or one of its pharmaceuticallyacceptable salts at an equivalent concentration.
 10. Formulationaccording to claim 1, obtained after freeze-drying a solution in whichthe mannitol is at a concentration of 10 mg per ml, the alanine is at aconcentration of 23 mg per ml and the active ingredient is irbesartan ata concentration of 1 mg per ml or one of its pharmaceutically acceptablesalts at an equivalent concentration.
 11. Formulation according to claim1, obtained after freeze-drying a solution in which the mannitol is at aconcentration of 9 mg per ml, the alanine is at a concentration of 18 mgper ml and the active ingredient is1-(2-naphthalen-2-ylethyl)-4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridinehydrochloride at a concentration of between 0.01 mg and 0.2 mg per ml orone of its pharmaceutically acceptable salts at an equivalentconcentration.
 12. Process for the stabilization of a nonprotein activeingredient contained in a freeze-dried pharmaceutically acceptableformulation, wherein the freeze-dried formulation is obtained startingfrom an aqueous solution comprising said active ingredient, mannitol andalanine in a ratio R of between 0.1 and 1, R representing the mass ofmannitol to the mass of alanine.
 13. Formulation according to claim 1,in which the active ingredient is selected from the group consisting ofphenylalkanoic acids, oxicam nonsteroid anti-inflammatory agents,paracetamol, lysine or arginine acetylsalicylate, bile acids,corticosteroids, anthracyclines, phloroglucinol, platinum derivatives,derivatives of alkaloids from Vinca minor, derivatives of alkaloids fromrye ergot, derivatives of purine or pyrimidine bases, prostaglandins,benzodiazepines, antibiotics, nitrosoureas, nitrogenous mustards, H₂antagonists, omeprazole, vitamins, antitumour agents, cardiovascularmedcines, haematological medicines, anticoagulant and antithromboticmedicine, heparinoids, diarginine oxoglutarate, plant extracts,nucleotides, valproic acid and its analogues, metopimazine, moxisylite,active bisphosphonates as antiosteoporotic agent, pralidoxime,deferoxamine, barbiturates, clomethiazole, 5-HT₂ antagonists,antagonists of angiotensin II, fantofarone, tirapazamine,(2S)-1-[(2R,3S)-5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzenesulphonyl)-3-hydroxy-2,3-dihydro-1H-indole-2-carbonyl]pyrrolidine-2-carboxamide,N,N-dibutyl-3-{4-[(2-butyl-5-methylsulphonamido)benzofuran-3-ylcarbonyl]phenoxyl}propylamine,6-(2-diethylamino-2-methyl)propylamino-3-phenyl-4-propylpyridazine,ethyl{(7S)-7-[(2R)-2-(e-chlorophenyl)-2-hydroxyethylamino]-5,6,7,8-tetrahydronaphthalen-2-yloxy}acetate,1-(2,4-dichlorophenyl)-3-(N-piperidin-1-ylcarboxamido)4-methyl-5-(4-chlorophenyl)-1H-pyrazole,4-{[N-(3,4-dimethoxyphenethyl)]-N-methylaminopropoxyl}-2-benzenesulphonyl-3-isopropyl-1-methylindole,2-{[1-(7-chloroquinolin-4-yl)-5-(2,6-dimethoxy-phenyl)-1H-pyrazole-3-carbonyl]amino}adamantane-2-carboxylicacid,(N-cyclohexyl-N-ethyl-3-(3-chloro-4-cyclohexylphenyl)prop-2-enylamine,(−)-N-methyl-N-[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl)butyl]benzamide,(S)-1-{2-[3-(3,4-dichlorophenyl)-1-(3-isopropoxyphenylacetyl)piperidin-3-yl]ethyl}-4-phenyl-1-azoniabicyclo[2.2.2]octanechloride and its pharmaceutically acceptable quaternary salts,4-amino-1-(6-chloropyrid-2-yl)piperidine,(S)-N-(1-{3-[1-benzoyl-3-(3,4-dichlorophenyl)piperidin-3-yl]propyl}4-phenylpiperidin4-yl)-N-methylacetamide,2-{[4-(2-chlorophenyl)thiazol-2-yl]aminocarbonyl}indole-1-acetic acid,clopidogrel,1-(2-naphthalen-2-ylethyl)4-(3-trifluoromethylphenyl)-1,2,3,6-tetrahydropyridinehydrochloride,N,N-dimethyl-N′-(pyridin-3-yl)methyl-N′-[4-(2,4,6-triisopropylphenyl)-thiazol-2-yl]ethane-1,2-diamine, and their pharmaceutically acceptable salts.